Can peritoneal B cells be rendered unresponsive?

Ly-1⁺ B cells have been reported to produce a number of autoantibodies, and to be involved in the selection and regulation of the conventional B cell repertoire. It is not known if these B cells, which are found in high numbers in the peritoneum of normal adult mice, themselves can be regulated. In this study, we evaluated the sensitivity of peritoneal B cells (PBCs) versus conventional splenic B cells to regulation in a model system for tolerance. Normal splenic (conventional) or PBCs (containing both CD5⁺ and CD5^- 'sister' cells) were cultured overnight with either F(ab')₂ or intact IgG anti-mouse Ig, washed, and then challenged with fluorescein(FL)-coupled to Brucella abortus (BA), trlmethylammonium (TMA)-BA or lipopolysaccharide (LPS), and the IgM responses to the FL and TMA haptens measured. In contrast to spleen cells, which exhibited up to a 90% reduction in anti-FL responsiveness, pretreated PBCs were mostly resistant to this form of tolerance regardless of challenge. The anti-TMA response of PBCs, which reflects the skewed V_H11 usage by peritoneal CD5 B cells, was also resistant to tolerance. However, spienic TMA-specific B cells appeared to be sensitive to unresponsiveness induced by anti-lg. Signaling studies show that PBCs have a blunted initial Ca²⁺ response, suggesting that the consequence of anti-lg crosslinklng may be defective in these cells. Furthermore, phorbal myristate acetate and/or lonomycln treatment of both PB and splenic B cells led to hyporesponsiveness to LPS challenge. This suggests that PBCs may be defective in a signalling pathway, perhaps involving protein klnase C activation. Our studies reveal that antibody production by PBCs is not subject to conventional tolerance pathways, a control which may be Important in terms of their ability to regulate other B cells.